National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Key Laboratory of Parasite and Vector Biology, National Health and Family Planning Commission, WHO Collaborating Center for Tropical Diseases, National Center for International Research on Tropical Diseases, Shanghai, 200025, People's Republic of China.
Joint Research Laboratory of Genetics and Ecology on Parasites-hosts Interaction, National Institute of Parasitic Diseases - Fudan University, Shanghai, 200025, China.
Parasit Vectors. 2018 Mar 12;11(1):172. doi: 10.1186/s13071-018-2734-7.
microRNAs (miRNAs) are small non-coding RNAs widely identified in many mosquitoes. They are reported to play important roles in development, differentiation and innate immunity. However, miRNAs in Anopheles sinensis, one of the Chinese malaria mosquitoes, remain largely unknown.
We investigated the global miRNA expression profile of An. sinensis using Illumina Hiseq 2000 sequencing. Meanwhile, we applied a bioinformatic approach to identify potential miRNAs in An. sinensis. The identified miRNA profiles were compared and analyzed by two approaches. The selected miRNAs from the sequencing result and the bioinformatic approach were confirmed with qRT-PCR. Moreover, target prediction, GO annotation and pathway analysis were carried out to understand the role of miRNAs in An. sinensis.
We identified 49 conserved miRNAs and 12 novel miRNAs by next-generation high-throughput sequencing technology. In contrast, 43 miRNAs were predicted by the bioinformatic approach, of which two were assigned as novel. Comparative analysis of miRNA profiles by two approaches showed that 21 miRNAs were shared between them. Twelve novel miRNAs did not match any known miRNAs of any organism, indicating that they are possibly species-specific. Forty miRNAs were found in many mosquito species, indicating that these miRNAs are evolutionally conserved and may have critical roles in the process of life. Both the selected known and novel miRNAs (asi-miR-281, asi-miR-184, asi-miR-14, asi-miR-nov5, asi-miR-nov4, asi-miR-9383, and asi-miR-2a) could be detected by quantitative real-time PCR (qRT-PCR) in the sequenced sample, and the expression patterns of these miRNAs measured by qRT-PCR were in concordance with the original miRNA sequencing data. The predicted targets for the known and the novel miRNAs covered many important biological roles and pathways indicating the diversity of miRNA functions. We also found 21 conserved miRNAs and eight counterparts of target immune pathway genes in An. sinensis based on the analysis of An. gambiae.
Our results provide the first lead to the elucidation of the miRNA profile in An. sinensis. Unveiling the roles of mosquito miRNAs will undoubtedly lead to a better understanding of mosquito biology and mosquito-pathogen interactions. This work lays the foundation for the further functional study of An. sinensis miRNAs and will facilitate their application in vector control.
微小 RNA（miRNAs）是在许多蚊子中广泛鉴定的小型非编码 RNA。据报道，它们在发育、分化和先天免疫中发挥重要作用。然而，中国疟疾蚊子之一的中华按蚊中的 miRNAs 仍知之甚少。
我们使用 Illumina Hiseq 2000 测序技术研究了中华按蚊的全局 miRNA 表达谱。同时，我们应用生物信息学方法鉴定了中华按蚊中的潜在 miRNAs。通过两种方法比较和分析鉴定的 miRNA 图谱。从测序结果和生物信息学方法中选择的 miRNA 通过 qRT-PCR 进行验证。此外，进行靶预测、GO 注释和途径分析，以了解 miRNA 在中华按蚊中的作用。
我们通过下一代高通量测序技术鉴定了 49 个保守 miRNA 和 12 个新 miRNA。相比之下，生物信息学方法预测了 43 个 miRNA，其中 2 个被指定为新 miRNA。通过两种方法对 miRNA 图谱的比较分析表明，它们之间有 21 个 miRNA 共享。12 个新 miRNA 与任何生物体的任何已知 miRNA 都不匹配，表明它们可能是物种特异性的。40 个 miRNA 在许多蚊子物种中发现，表明这些 miRNA 在进化上是保守的，并且可能在生命过程中具有关键作用。通过定量实时 PCR（qRT-PCR）在测序样本中可以检测到所选的已知和新 miRNA（asi-miR-281、asi-miR-184、asi-miR-14、asi-miR-nov5、asi-miR-nov4、asi-miR-9383 和 asi-miR-2a），并且通过 qRT-PCR 测量的这些 miRNA 的表达模式与原始 miRNA 测序数据一致。已知和新 miRNA 的预测靶标涵盖了许多重要的生物学作用和途径，表明 miRNA 功能的多样性。我们还基于对冈比亚按蚊的分析，在中华按蚊中发现了 21 个保守 miRNA 和 8 个免疫途径基因的对应物。
我们的结果为阐明中华按蚊的 miRNA 图谱提供了第一个线索。揭示蚊子 miRNA 的作用无疑将导致对蚊子生物学和蚊子-病原体相互作用的更好理解。这项工作为中华按蚊 miRNA 的进一步功能研究奠定了基础，并将促进它们在媒介控制中的应用。
